TY - JOUR

T1 - RPA mediates recombination repair during replication stress and is displaced from DNA by checkpoint signalling in human cells.

AU - Sleeth, Kate M

AU - Sørensen, Claus Storgaard

AU - Issaeva, Natalia

AU - Dziegielewski, Jaroslaw

AU - Bartek, Jiri

AU - Helleday, Thomas

N1 - Keywords: Cell Line, Tumor; DNA Damage; DNA Repair; DNA Replication; Enzyme Activation; Genes, cdc; Humans; Hydroxyurea; Nucleic Acid Synthesis Inhibitors; Protein Kinases; Rad51 Recombinase; Rad52 DNA Repair and Recombination Protein; Replication Protein A; Signal Transduction

PY - 2007

Y1 - 2007

N2 - The replication protein A (RPA) is involved in most, if not all, nuclear metabolism involving single-stranded DNA. Here, we show that RPA is involved in genome maintenance at stalled replication forks by the homologous recombination repair system in humans. Depletion of the RPA protein inhibited the formation of RAD51 nuclear foci after hydroxyurea-induced replication stalling leading to persistent unrepaired DNA double-strand breaks (DSBs). We demonstrate a direct role of RPA in homology directed recombination repair. We find that RPA is dispensable for checkpoint kinase 1 (Chk1) activation and that RPA directly binds RAD52 upon replication stress, suggesting a direct role in recombination repair. In addition we show that inhibition of Chk1 with UCN-01 decreases dissociation of RPA from the chromatin and inhibits association of RAD51 and RAD52 with DNA. Altogether, our data suggest a direct role of RPA in homologous recombination in assembly of the RAD51 and RAD52 proteins. Furthermore, our data suggest that replacement of RPA with the RAD51 and RAD52 proteins is affected by checkpoint signalling.

AB - The replication protein A (RPA) is involved in most, if not all, nuclear metabolism involving single-stranded DNA. Here, we show that RPA is involved in genome maintenance at stalled replication forks by the homologous recombination repair system in humans. Depletion of the RPA protein inhibited the formation of RAD51 nuclear foci after hydroxyurea-induced replication stalling leading to persistent unrepaired DNA double-strand breaks (DSBs). We demonstrate a direct role of RPA in homology directed recombination repair. We find that RPA is dispensable for checkpoint kinase 1 (Chk1) activation and that RPA directly binds RAD52 upon replication stress, suggesting a direct role in recombination repair. In addition we show that inhibition of Chk1 with UCN-01 decreases dissociation of RPA from the chromatin and inhibits association of RAD51 and RAD52 with DNA. Altogether, our data suggest a direct role of RPA in homologous recombination in assembly of the RAD51 and RAD52 proteins. Furthermore, our data suggest that replacement of RPA with the RAD51 and RAD52 proteins is affected by checkpoint signalling.

U2 - 10.1016/j.jmb.2007.07.068

DO - 10.1016/j.jmb.2007.07.068

M3 - Journal article

C2 - 17765923

VL - 373

SP - 38

EP - 47

JO - Journal of Molecular Biology

JF - Journal of Molecular Biology

SN - 0022-2836

IS - 1

ER -